Role of ankyrin-B in human arrhythmia

描述（由申请人提供）：心脏兴奋性缺陷是人类心律失常和心源性猝死的基础，是发达国家死亡的主要原因。离子通道和转运体控制着带电离子穿过细胞膜的运动。在心脏中，这些蛋白质的协调活动调节跨膜电化学梯度来控制去极化/再极化，从而控制心脏的兴奋性。离子通道和转运体的正常功能需要明确的生物物理特性，以及在明确的膜域中精确的表达、组织和调节。在我们第一期资助期间产生的发现支持了人类心脏病（心律失常）的新范式，该范式基于特定可兴奋膜上离子通道和转运体的适当表达和局部调节所需的蛋白质功能障碍。具体来说，我们发现锚蛋白，以前被认为是静态膜适配器，在心室心肌细胞的离子通道，转运体和信号蛋白靶向中发挥动态作用。锚蛋白b基因（ANK2）功能缺失突变的患者表现出严重而复杂的心脏表型。表型可能包括窦房结功能障碍、心房颤动（AF）、传导缺陷、儿茶酚胺诱导的多形性室性心律失常和/或心源性猝死。此外，我们已经了解到，普通人群中常见的ANK2基因变异与QTc改变和室性心律失常易感性相关，AnkB水平在心血管疾病的大型动物模型中发生强烈改变，并且ANK2是普通人群中AF易感性的候选基因。然而，尽管这些翻译研究表明AnkB在心脏兴奋性中起着关键作用，但令人惊讶的是，AnkB在心脏中的具体分子作用仍然未知。事实上，ankb靶向途径（或其他心脏靶向途径）的体内细胞成分的身份仍然未知。最后，缺乏AnkB缺失的动物模型（全局AnkB k/o是胚胎致死的）阻碍了确定AnkB在心脏生理和疾病中的新作用的努力。对于这第一次竞争性更新，由于第一个融资周期的重要进展和许多创新的新动物模型的开发，我们处于有利地位，可以在基线和疾病中提供关于整个ankb靶向途径的基本成分（包括上游和下游）的第一个体内信息。我们提供了令人兴奋的新初步数据，确定了一个调节心膜兴奋性并与AnkB相关的膜转运蛋白（EHD蛋白）的新家族。我们进一步提供了新的数据，表明AnkB在窦房结和心房的选择性Ca2+通道中起着新的作用。最后，我们在小鼠身上的初步数据显示了AnkB在心膜生物发生和维持中的新的和意想不到的作用。总之，我们发表的研究结果和初步数据支持一个中心假设，即基于ankb的细胞通路在肌细胞膜兴奋性和心功能中起动态作用。我们的研究计划的直接目标是了解AnkB在心脏中的特定细胞作用（包括上游调控途径[EHD蛋白]和新的下游靶点[Cav1.3]），并确定AnkB功能障碍如何导致复杂的人类心脏病。对于这第一次竞争性更新，我们提出了一系列未表征和创新的动物模型，新的分子工具，创新技术和新的抗体来测试AnkB细胞途径在体内的特定作用。

DESCRIPTION (provided by applicant): Defects in cardiac excitability are the basis for human arrhythmia and sudden cardiac death, a leading cause of mortality in developed countries. Ion channels and transporters control the movement of charged ions across cell membranes. In the heart, the coordinate activities of these proteins regulate the transmembrane electrochemical gradient to control depolarization/repolarization, and thus cardiac excitability. Normal function of ion channels and transporters requires defined biophysical properties as well as precise expression, organization, and regulation in defined membrane domains. Findings generated during our first period of funding support a new paradigm for human cardiac disease (arrhythmia) based on dysfunction in proteins that are required for proper expression and local regulation of ion channels and transporters at specific excitable membranes. Specifically, we discovered that ankyrin proteins, previously considered static membrane adapters, play dynamic roles in ion channel, transporter, and signaling protein targeting in ventricular cardiomyocytes. Patients harboring loss-of-function mutations in the ankyrin-B gene (ANK2) display a severe and complex cardiac phenotype. Phenotypes may include sinus node dysfunction, atrial fibrillation (AF), conduction defects, catecholamine-induced polymorphic ventricular arrhythmia, and/or sudden cardiac death. Moreover, we have learned that common ANK2 gene variants in the general population are associated with QTc alterations and ventricular arrhythmia susceptibility, that AnkB levels are strongly altered in large animal models of cardiovascular disease, and that the ANK2 is a candidate gene for AF susceptibility in the general human population. However, despite these translational studies implicating AnkB as a key player in cardiac excitability, the specific molecular roles of AnkB in heart remain surprisingly unknown. In fact, the identities of the in vivo cellular components of the AnkB-targeting pathway (or other cardiac targeting pathways) are still unknown. Finally, lack of an animal model of AnkB deficiency (global AnkB k/o is embryonic lethal) has prevented efforts to define new roles of AnkB in cardiac physiology and disease. For this first competitive renewal, due to important advances during the first funding cycle and the development of a number of innovative new animal models, we are well-positioned to provide the first in vivo information on the fundamental components (both upstream & downstream) of the entire AnkB-targeting pathway at baseline and in disease. We provide exciting new preliminary data that identifies a novel family of membrane trafficking proteins (EHD proteins) that regulate cardiac membrane excitability and associate with AnkB. We further provide new data that AnkB plays a novel role in targeting select Ca2+ channels in sinus node & atria. Finally, our preliminary data in mice demonstrates novel and unexpected roles of AnkB in cardiac membrane biogenesis and maintenance. Together, our published findings and preliminary data support a central hypothesis that the AnkB-based cellular pathway plays dynamic roles in myocyte membrane excitability and cardiac function. The immediate goals of our research program are to understand the specific cellular role(s) of AnkB in the heart (including upstream regulatory pathways [EHD proteins] and novel downstream targets [Cav1.3]) and determine how AnkB dysfunction leads to complex human cardiac disease. For this first competitive renewal, we present a cast of uncharacterized and innovative animal models, novel molecular tools, innovative technologies, and new antibodies to test the specific roles of the AnkB cellular pathway in vivo.